The bacterium Escherichia coli can survive in a wide variety of environments due to its ability to rapidly adapt to changing conditions via a repertoire of stress responses. One type of stress is an increase in the osmotic strength of the growth medium; the ability to adapt is relevant to the bacterium's survival in the human bladder and colon, sewage systems, and seawater. Adaptive alterations in cellular metabolism and structure are mediated by the products of certain osmotically responsive genes. Our primary goal is to understand the molecular mechanisms governing osmotically dependent gene expression. An excellent experimental model is proU, a locus which encodes a transport system for the ubiquitous osmoprotectant glycine betaine. proU is induced strongly and rapidly in response to the changes in intracellular ion composition that accompany osmotic stress. Transcription activation in vitro requires only the DNA template, RNA polymerase and potassium glutamate. Genetic manipulation of both the promoter and RNA polymerase and biochemical dissection of events effected by potassium glutamate will provide the molecular details of this unique type of transcriptional control. The analysis will be extended to other genes responding to osmotic signals to determine whether there is a common regulatory mechanism. osmb, which encodes a structural lipoprotein, is representative of a class of genes that are induced by two signals, growth in hyperosmotic media and stationary phase. In vitro gene expression will determine the signal transduction mechanism for each of these stimuli and, along with genetic approaches, identify necessary trans-acting regulatory factors.